Diabetes mellitus is a chronic disorder of carbohydrate metabolism characterized by insufficient production of insulin by the pancreatic beta cells. Diabetes effects approximately 10 million people in the United States, with more than 250,000 new cases diagnosed each year. There are two common types of diabetes mellitus: insulin-dependent (Type-I diabetes) and non-insulin-dependent (Type-II diabetes). Insulin-dependent diabetes is generally characterized by an absolute deficiency of insulin production, whereas non-insulin-dependent diabetes is characterized by a relatively insufficient production of insulin.
In normal individuals, the rate of insulin secretion by beta cells is regulated by the level of glucose in the blood. When the blood glucose level rises, the islet cells are stimulated to release increased amounts of insulin into the blood, accelerating glucose transport into the cells and glucose conversion into glycogen. As the blood glucose level falls, insulin release from the islets is decreased. In the diabetic subject, insulin production is abnormally low or insufficient, resulting in abnormally high blood glucose levels, a condition known as hyperglycemia.
In addition to diet and exercise programs, the constant and life long monitoring of blood glucose levels in conjunction with injections of insulin is central to the current methods for the treatment of the insulin-dependent diabetic subject. Many diabetic subjects, however, have difficulty in controlling their blood glucose levels using the current treatment methods, thus constantly exposing themselves to the adverse effects of hypoglycemia (abnormally low blood glucose levels) and hyperglycemia.
The inability to precisely control the level of blood glucose also poses long term complications such as degenerative vascular changes (e.g. atherosclerosis and microangiopathy), neuropathy (e.g. peripheral nerve degeneration, autonomic nervous system, and cranial nerve lesions), ocular disturbances (e.g. blurred vision, cataracts, and diabetic retinopathy), kidney diseases (e.g. recurrent pyelonephritis and nephropathy), and infections. Accordingly, there exists a need for an alternative method for controlling blood glucose levels in the diabetic patient. The transplantation of beta cells has been proposed as an alternative therapy in the treatment of diabetes. However, large scale transplantation of human beta cells is not feasible because of the limited availability of donors; similarly, the cost and effort, in terms of labor, associated with obtaining sufficient amounts of animal islets for transplantation also limits their use. These and other disadvantages associated with transplanting human and animal islets, makes the development of islet-derived cell lines the method of choice in obtaining sufficient quantities of cells for transplantation. In particular, a number of beta cell lines have been generated from insulinomas and hyperplastic islets arising in mice expressing a transgene encoding the SV40 T antigen (Tag) oncogene under the control of the insulin promoter (RIP-Tag)(1-6). Several of these cell lines displayed insulin secretion characteristics similar to those observed in intact adult islets, in particular the response to glucose concentrations in the physiological range (5-15 mmol/l).
However, a common problem encountered with all of these cell lines is their phenotypic instability. After propagation in tissue culture, these cells become responsive to subphysiological concentrations of glucose and/or manifest diminished insulin output (4, 6-9). A similar instability has been observed with beta cell lines derived by other methods (10-12).
The present invention overcomes the problems associated with the previous beta cell lines by providing a beta cell line which not only maintains blood glucose levels in the normal range, but also may be controlled to prevent unregulated proliferation.